Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 81
Filtrar
Mais filtros

Base de dados
País/Região como assunto
Tipo de documento
Intervalo de ano de publicação
1.
Cell ; 174(5): 1200-1215.e20, 2018 08 23.
Artigo em Inglês | MEDLINE | ID: mdl-30100187

RESUMO

Nuclear pore complexes (NPCs) regulate nuclear-cytoplasmic transport, transcription, and genome integrity in eukaryotic cells. However, their functional roles in cancer remain poorly understood. We interrogated the evolutionary transcriptomic landscape of NPC components, nucleoporins (Nups), from primary to advanced metastatic human prostate cancer (PC). Focused loss-of-function genetic screen of top-upregulated Nups in aggressive PC models identified POM121 as a key contributor to PC aggressiveness. Mechanistically, POM121 promoted PC progression by enhancing importin-dependent nuclear transport of key oncogenic (E2F1, MYC) and PC-specific (AR-GATA2) transcription factors, uncovering a pharmacologically targetable axis that, when inhibited, decreased tumor growth, restored standard therapy efficacy, and improved survival in patient-derived pre-clinical models. Our studies molecularly establish a role of NPCs in PC progression and give a rationale for NPC-regulated nuclear import targeting as a therapeutic strategy for lethal PC. These findings may have implications for understanding how NPC deregulation contributes to the pathogenesis of other tumor types.


Assuntos
Fator de Transcrição E2F1/metabolismo , Glicoproteínas de Membrana/metabolismo , Poro Nuclear/fisiologia , Neoplasias da Próstata/metabolismo , Proteínas Proto-Oncogênicas c-myc/metabolismo , Fatores de Transcrição/metabolismo , Transporte Ativo do Núcleo Celular , Carcinogênese , Núcleo Celular/metabolismo , Proliferação de Células , Fator de Transcrição GATA2/metabolismo , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Membrana Nuclear , Complexo de Proteínas Formadoras de Poros Nucleares , Transdução de Sinais
2.
BMC Bioinformatics ; 21(1): 56, 2020 Feb 13.
Artigo em Inglês | MEDLINE | ID: mdl-32054449

RESUMO

BACKGROUND: Quality Control in any high-throughput sequencing technology is a critical step, which if overlooked can compromise an experiment and the resulting conclusions. A number of methods exist to identify biases during sequencing or alignment, yet not many tools exist to interpret biases due to outliers. RESULTS: Hence, we developed iSeqQC, an expression-based QC tool that detects outliers either produced due to variable laboratory conditions or due to dissimilarity within a phenotypic group. iSeqQC implements various statistical approaches including unsupervised clustering, agglomerative hierarchical clustering and correlation coefficients to provide insight into outliers. It can be utilized through command-line (Github: https://github.com/gkumar09/iSeqQC) or web-interface (http://cancerwebpa.jefferson.edu/iSeqQC). A local shiny installation can also be obtained from github (https://github.com/gkumar09/iSeqQC). CONCLUSION: iSeqQC is a fast, light-weight, expression-based QC tool that detects outliers by implementing various statistical approaches.


Assuntos
Perfilação da Expressão Gênica/normas , Sequenciamento de Nucleotídeos em Larga Escala/normas , Análise de Sequência de RNA/normas , Software , Análise por Conglomerados , Humanos , Controle de Qualidade
3.
J Immunol ; 200(3): 1133-1145, 2018 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-29288198

RESUMO

Recent work indicates that salivary glands are able to constitutively recruit CD8+ T cells and retain them as tissue-resident memory T cells, independently of local infection, inflammation, or Ag. To understand the mechanisms supporting T cell recruitment to the salivary gland, we compared T cell migration to the salivary gland in mice that were infected or not with murine CMV (MCMV), a herpesvirus that infects the salivary gland and promotes the accumulation of salivary gland tissue-resident memory T cells. We found that acute MCMV infection increased rapid T cell recruitment to the salivary gland but that equal numbers of activated CD8+ T cells eventually accumulated in infected and uninfected glands. T cell recruitment to uninfected salivary glands depended on chemokines and the integrin α4 Several chemokines were expressed in the salivary glands of infected and uninfected mice, and many of these could promote the migration of MCMV-specific T cells in vitro. MCMV infection increased the expression of chemokines that interact with the receptors CXCR3 and CCR5, but neither receptor was needed for T cell recruitment to the salivary gland during MCMV infection. Unexpectedly, however, the chemokine receptor CXCR3 was critical for T cell accumulation in uninfected salivary glands. Together, these data suggest that CXCR3 and the integrin α4 mediate T cell recruitment to uninfected salivary glands but that redundant mechanisms mediate T cell recruitment after MCMV infection.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Infecções por Herpesviridae/imunologia , Integrina alfa4/genética , Muromegalovirus/imunologia , Receptores CXCR3/genética , Glândulas Salivares/imunologia , Animais , Movimento Celular/imunologia , Células Cultivadas , Quimiocinas/metabolismo , Infecções por Herpesviridae/virologia , Memória Imunológica/imunologia , Interferon gama/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptores CCR5/genética , Glândulas Salivares/virologia
4.
J Cell Physiol ; 234(8): 13042-13056, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-30552679

RESUMO

We previously reported that HOXA4, HOXA9, and HOXD10 are selectively expressed in colonic stem cells (SCs) and their overexpression contributes to colorectal cancer (CRC). Our goals here were to determine how these HOX genes are transcriptionally regulated and whether transcriptional dysregulation of HOX genes occurs in CRC. Accordingly, we used correlation analysis to identify genes that are expression-correlated or anticorrelated with HOXA4, HOXA9, and HOXD10. We then used Gene Ontology (GO) analysis to functionally classify these genes. The GO results for both HOXA4 and HOXD10 correlated gene sets for normal colon and CRC show functions mostly classified as developmental, transcriptional regulation, and DNA binding. This raised the question: Are these gene sets regulated by the same transcription factors (TFs)? Consequently, we used promoter analysis and interaction network toolset (PAINT) to identify commonly shared transcription response elements. The results indicated that completely different sets of TFs coregulate HOXA4 and HOXD10 (but not HOXA9) and their expression-correlated genes. And predicted TFs are altered in CRC compared with normal colon. Taken together, analysis of gene signatures correlated with expression of HOXA4 and HOXD10 indicates how these HOX genes are: (a) transcriptionally regulated in the normal colon; (b) dysregulated in CRC. This discovery provides a mechanism for targeting CRC SCs.


Assuntos
Neoplasias do Colo/genética , Regulação Neoplásica da Expressão Gênica/fisiologia , Proteínas de Homeodomínio/biossíntese , Fatores de Transcrição/biossíntese , Neoplasias do Colo/patologia , Redes Reguladoras de Genes , Proteínas de Homeodomínio/genética , Humanos , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Transcriptoma
5.
Mol Cell ; 43(4): 663-72, 2011 Aug 19.
Artigo em Inglês | MEDLINE | ID: mdl-21855804

RESUMO

The LXCXE peptide motif facilitates interaction between the RB tumor suppressor and a large number of cellular proteins that are expected to impinge on diverse biological processes. In vitro and in vivo analyses demonstrated that LXCXE binding function is dispensable for RB promoter association and control of basal gene expression. Dependence on this function of RB is unmasked after DNA damage, wherein LXCXE binding is essential for exerting control over E2F3 and suppressing cell-cycle progression in the presence of genotoxic stress. Gene expression profiling revealed that the transcriptional program coordinated by this specific aspect of RB is associated with progression of human hepatocellular carcinoma and poor disease outcome. Consistent with these findings, biological challenge revealed a requirement for LXCXE binding in suppression of genotoxin-initiated hepatocellular carcinoma in vivo. Together, these studies establish an essential role of the LXCXE binding motif for RB-mediated transcriptional control, response to genotoxic insult, and tumor suppression.


Assuntos
Dano ao DNA , Regulação da Expressão Gênica , Proteína do Retinoblastoma/fisiologia , Transcrição Gênica , Motivos de Aminoácidos , Animais , Sítios de Ligação , Linhagem Celular , Cromatina/metabolismo , Humanos , Camundongos , Domínios e Motivos de Interação entre Proteínas , Mapeamento de Interação de Proteínas , Proteína do Retinoblastoma/química , Proteína do Retinoblastoma/metabolismo
6.
Hepatology ; 60(4): 1231-40, 2014 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-24824777

RESUMO

UNLABELLED: Cancers mediated by viral etiology must exhibit deregulated cellular proliferation and evade immune recognition. The role of the retinoblastoma tumor suppressor (RB) pathway, which is lost at relatively high frequency in hepatocellular carcinoma (HCC), has recently been expanded to include the regulation of innate immune responsiveness. In this study we investigated the coordinate impact of RB-loss on cell cycle control and immune function in the liver. We found that RB depletion in hepatoma cells resulted in a compromised immunological response to multiple stimuli and reduced the potential of these cells to recruit myeloid cells. Viral-mediated liver-specific RB deletion in vivo led to the induction of genes associated with proliferation and cell cycle entry as well as the significant attenuation of genes associated with immune function, as evidenced by decreases in cytokine and chemokine expression, leukocyte recruitment, and hepatic inflammation. To determine if these changes in gene expression were instructive in human disease, we compared our liver-specific RB-loss gene signature to existing profiles of HCC and found that this signature was associated with disease progression and confers a worse prognosis. CONCLUSION: Our data confirm that RB participates in the regulation of innate immunity in liver parenchymal cells both in vitro and in vivo and to our knowledge describes the first gene signature associated with HCC that includes both immunoregulatory and proliferative genes and that can also be attributed to the alteration of a single gene in vitro.


Assuntos
Carcinoma Hepatocelular/fisiopatologia , Hepatócitos/imunologia , Hepatócitos/patologia , Imunidade Inata/fisiologia , Neoplasias Hepáticas/fisiopatologia , Proteína do Retinoblastoma/fisiologia , Adenoviridae , Animais , Carcinoma Hepatocelular/imunologia , Carcinoma Hepatocelular/patologia , Ciclo Celular/fisiologia , Linhagem Celular Tumoral , Proliferação de Células/fisiologia , Citocinas/metabolismo , Progressão da Doença , Humanos , Técnicas In Vitro , Neoplasias Hepáticas/imunologia , Neoplasias Hepáticas/patologia , Masculino , Camundongos , Camundongos Mutantes , Proteína do Retinoblastoma/genética , Transfecção
7.
Am J Pathol ; 182(6): 2264-75, 2013 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-23660011

RESUMO

The molecular mechanisms underlying progression of prostate cancer (PCa) to castrate-resistant (CR) and metastatic disease are poorly understood. Our previous mechanistic work shows that inhibition of transcription factor Stat5 by multiple alternative methods induces extensive rapid apoptotic death of Stat5-positive PCa cells in vitro and inhibits PCa xenograft tumor growth in nude mice. Furthermore, STAT5A/B induces invasive behavior of PCa cells in vitro and in vivo, suggesting involvement of STAT5A/B in PCa progression. Nuclear STAT5A/B protein levels are increased in high-grade PCas, CR PCas, and distant metastases, and high nuclear STAT5A/B expression predicts early disease recurrence and PCa-specific death in clinical PCas. Based on these findings, STAT5A/B represents a therapeutic target protein for advanced PCa. The mechanisms underlying increased Stat5 protein levels in PCa are unclear. Herein, we demonstrate amplification at the STAT5A/B gene locus in a significant fraction of clinical PCa specimens. STAT5A/B gene amplification was more frequently found in PCas of high histologic grades and in CR distant metastases. Quantitative in situ analysis revealed that STAT5A/B gene amplification was associated with increased STAT5A/B protein expression in PCa. Functional studies showed that increased STAT5A/B copy numbers conferred growth advantage in PCa cells in vitro and as xenograft tumors in vivo. The work presented herein provides the first evidence of somatic STAT5A/B gene amplification in clinical PCas.


Assuntos
Amplificação de Genes , Neoplasias da Próstata/genética , Fator de Transcrição STAT5/genética , Proteínas Supressoras de Tumor/genética , Animais , Variações do Número de Cópias de DNA , DNA de Neoplasias/genética , Progressão da Doença , Regulação Neoplásica da Expressão Gênica/fisiologia , Humanos , Hibridização in Situ Fluorescente , Masculino , Camundongos , Camundongos Nus , Gradação de Tumores , Transplante de Neoplasias , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Recidiva , Fator de Transcrição STAT5/biossíntese , Transplante Heterólogo , Células Tumorais Cultivadas , Proteínas Supressoras de Tumor/biossíntese
8.
BMC Genomics ; 14: 1, 2013 Jan 16.
Artigo em Inglês | MEDLINE | ID: mdl-23323973

RESUMO

BACKGROUND: Human blood platelets are essential to maintaining normal hemostasis, and platelet dysfunction often causes bleeding or thrombosis. Estimates of genome-wide platelet RNA expression using microarrays have provided insights to the platelet transcriptome but were limited by the number of known transcripts. The goal of this effort was to deep-sequence RNA from leukocyte-depleted platelets to capture the complex profile of all expressed transcripts. RESULTS: From each of four healthy individuals we generated long RNA (≥40 nucleotides) profiles from total and ribosomal-RNA depleted RNA preparations, as well as short RNA (<40 nucleotides) profiles. Analysis of ~1 billion reads revealed that coding and non-coding platelet transcripts span a very wide dynamic range (≥16 PCR cycles beyond ß-actin), a result we validated through qRT-PCR on many dozens of platelet messenger RNAs. Surprisingly, ribosomal-RNA depletion significantly and adversely affected estimates of the relative abundance of transcripts. Of the known protein-coding loci, ~9,500 are present in human platelets. We observed a strong correlation between mRNAs identified by RNA-seq and microarray for well-expressed mRNAs, but RNASeq identified many more transcripts of lower abundance and permitted discovery of novel transcripts. CONCLUSIONS: Our analyses revealed diverse classes of non-coding RNAs, including: pervasive antisense transcripts to protein-coding loci; numerous, previously unreported and abundant microRNAs; retrotransposons; and thousands of novel un-annotated long and short intronic transcripts, an intriguing finding considering the anucleate nature of platelets. The data are available through a local mirror of the UCSC genome browser and can be accessed at: http://cm.jefferson.edu/platelets_2012/.


Assuntos
Plaquetas/citologia , Plaquetas/metabolismo , Núcleo Celular , Genômica , Transcrição Gênica , Mineração de Dados , Humanos , Internet , Íntrons/genética , Pseudogenes/genética , RNA Antissenso/genética , RNA Mensageiro/genética , RNA Ribossômico/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Análise de Sequência de RNA
9.
Mol Cancer ; 12: 59, 2013 Jun 12.
Artigo em Inglês | MEDLINE | ID: mdl-23758962

RESUMO

BACKGROUND: Prolactin (PRL) is essential for normal mammary gland development. PRL promotes mammary tumor formation in rodents and elevated serum prolactin is associated with increased risk of estrogen-receptor positive breast cancer in women. On the other hand, PRL may also exert pro-differentiation effects and act to suppress invasive features of established breast cancer. Previously published limited global transcript profiling analyses of prolactin-regulated gene expression in human breast cancer cells have exclusively been performed in vitro. The present study aimed to shed new light on how PRL modulates estrogen receptor (ER)-positive breast cancer through global transcript profiling of a human breast cancer xenograft model in vivo. METHODS: The prolactin-responsive human T47D breast cancer cell line was xenotransplanted into nude mice and global transcript profiling was carried out following treatment with or without human PRL for 48 h. A subset of PRL-modulated transcripts was further validated using qRT-PCR and immunohistochemistry. RESULTS: The in vivo analyses identified 130 PRL-modulated transcripts, 75 upregulated and 55 downregulated, based on fold change >1.6 and P-value <0.05. From this initial panel of transcripts, a subset of 18 transcripts with established breast cancer-relevance were selected and validated by qRT-PCR. Some but not all of the transcripts were also PRL-modulated in vitro. The selected PRL-modulated transcripts were tested for dependence on Stat5, Jak1 or Jak2 activation, and for co-regulation by 17ß-estradiol (E2). The protein encoded by one of the PRL-regulated transcripts, PTHrP, was examined in a panel of 92 human breast cancers and found by in situ quantitative immunofluorescence analysis to be highly positively correlated with nuclear localized and tyrosine phosphorylated Stat5. Gene Ontology analysis revealed that PRL-upregulated genes were enriched in pathways involved in differentiation. Finally, a gene signature based on PRL-upregulated genes was associated with prolonged relapse-free and metastasis-free survival in breast cancer patients. CONCLUSIONS: This global analysis identified and validated a panel of PRL-modulated transcripts in an ER-positive human breast cancer xenotransplant model, which may have value as markers of relapse-free and metastasis-free survival. Gene products identified in the present study may facilitate ongoing deciphering of the pleiotropic effects of PRL on human breast cancer.


Assuntos
Neoplasias da Mama/genética , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Prolactina/farmacologia , Animais , Biomarcadores Tumorais/metabolismo , Neoplasias da Mama/patologia , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/genética , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/genética , Estradiol/farmacologia , Feminino , Ontologia Genética , Humanos , Janus Quinase 1/metabolismo , Janus Quinase 2/metabolismo , Camundongos Nus , Metástase Neoplásica , Análise de Sequência com Séries de Oligonucleotídeos , Proteína Relacionada ao Hormônio Paratireóideo/metabolismo , Fosfotirosina/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Reprodutibilidade dos Testes , Fator de Transcrição STAT5/metabolismo , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/genética , Ensaios Antitumorais Modelo de Xenoenxerto
10.
J Virol ; 86(6): 3200-10, 2012 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-22238315

RESUMO

Postexposure treatment (PET) of wild-type rabies virus (RV)-infected mice with a live-attenuated triple-glycoprotein RV variant (TriGAS) promotes survival but does not prevent the pathogenic RV from invading and replicating in the brain. Successful PET is associated with the induction of a robust virus-neutralizing antibody response and clearance of the wild-type RV from brain tissues. Comparison of the transcriptomes of normal mouse brain with those of wild-type-RV-infected mice that had received either mock or TriGAS PET treatment revealed that many of the host genes activated in the mock-treated mice represent type I interferon (IFN) response genes. This indicates that RV infection induces an early type I IFN response that is unable to control the infection. In contrast, most of the activated genes in the brain of the RV-infected, TriGAS-treated mouse play a role in adaptive immunity, including the regulation of T cell activation, T cell differentiation, and the regulation of lymphocyte and mononuclear cell proliferation. These findings were confirmed by quantitative PCR (qPCR) array studies, which showed that 3 genes in particular, encoding chemokine ligand 3 (Ccl3), natural killer cell activator 2 (interleukin 12B [IL-12B]), and granzyme A (GzmA), were activated earlier and to a greater extent in the brains of RV-infected mice treated with TriGAS than in the brains of mock-treated mice. The activation of these genes, known to play key roles in the regulation of lymphocyte and mononuclear cell proliferation, is likely an important part of the mechanism by which TriGAS mediates its PET activity.


Assuntos
Imunidade Adaptativa , Sistema Nervoso Central/imunologia , Vacina Antirrábica/imunologia , Vírus da Raiva/imunologia , Raiva/genética , Raiva/prevenção & controle , Regulação para Cima , Animais , Anticorpos Antivirais/imunologia , Sistema Nervoso Central/virologia , Feminino , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Profilaxia Pós-Exposição , Raiva/tratamento farmacológico , Raiva/imunologia , Vacina Antirrábica/uso terapêutico , Vacinas Atenuadas/imunologia , Vacinas Atenuadas/uso terapêutico
11.
Cell Rep Med ; 4(2): 100937, 2023 02 21.
Artigo em Inglês | MEDLINE | ID: mdl-36787737

RESUMO

Metastatic prostate cancer (PCa) inevitably acquires resistance to standard therapy preceding lethality. Here, we unveil a chromosomal instability (CIN) tolerance mechanism as a therapeutic vulnerability of therapy-refractory lethal PCa. Through genomic and transcriptomic analysis of patient datasets, we find that castration and chemotherapy-resistant tumors display the highest CIN and mitotic kinase levels. Functional genomics screening coupled with quantitative phosphoproteomics identify MASTL kinase as a survival vulnerability specific of chemotherapy-resistant PCa cells. Mechanistically, MASTL upregulation is driven by transcriptional rewiring mechanisms involving the non-canonical transcription factors androgen receptor splice variant 7 and E2F7 in a circuitry that restrains deleterious CIN and prevents cell death selectively in metastatic therapy-resistant PCa cells. Notably, MASTL pharmacological inhibition re-sensitizes tumors to standard therapy and improves survival of pre-clinical models. These results uncover a targetable mechanism promoting high CIN adaptation and survival of lethal PCa.


Assuntos
Neoplasias de Próstata Resistentes à Castração , Masculino , Humanos , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Neoplasias de Próstata Resistentes à Castração/metabolismo , Neoplasias de Próstata Resistentes à Castração/patologia , Receptores Androgênicos/genética , Receptores Androgênicos/metabolismo , Instabilidade Cromossômica , Proteínas Associadas aos Microtúbulos/genética , Proteínas Associadas aos Microtúbulos/uso terapêutico , Proteínas Serina-Treonina Quinases/genética
12.
Cancer Discov ; 13(12): 2584-2609, 2023 12 12.
Artigo em Inglês | MEDLINE | ID: mdl-37676710

RESUMO

Signaling rewiring allows tumors to survive therapy. Here we show that the decrease of the master regulator microphthalmia transcription factor (MITF) in lethal prostate cancer unleashes eukaryotic initiation factor 3B (eIF3B)-dependent translation reprogramming of key mRNAs conferring resistance to androgen deprivation therapy (ADT) and promoting immune evasion. Mechanistically, MITF represses through direct promoter binding eIF3B, which in turn regulates the translation of specific mRNAs. Genome-wide eIF3B enhanced cross-linking immunoprecipitation sequencing (eCLIP-seq) showed specialized binding to a UC-rich motif present in subsets of 5' untranslated regions. Indeed, translation of the androgen receptor and major histocompatibility complex I (MHC-I) through this motif is sensitive to eIF3B amount. Notably, pharmacologic targeting of eIF3B-dependent translation in preclinical models sensitizes prostate cancer to ADT and anti-PD-1 therapy. These findings uncover a hidden connection between transcriptional and translational rewiring promoting therapy-refractory lethal prostate cancer and provide a druggable mechanism that may transcend into effective combined therapeutic strategies. SIGNIFICANCE: Our study shows that specialized eIF3B-dependent translation of specific mRNAs released upon downregulation of the master transcription factor MITF confers castration resistance and immune evasion in lethal prostate cancer. Pharmacologic targeting of this mechanism delays castration resistance and increases immune-checkpoint efficacy. This article is featured in Selected Articles from This Issue, p. 2489.


Assuntos
Neoplasias de Próstata Resistentes à Castração , Neoplasias da Próstata , Masculino , Humanos , Fatores de Transcrição , Antagonistas de Androgênios/farmacologia , Antagonistas de Androgênios/uso terapêutico , Evasão da Resposta Imune , Receptores Androgênicos/genética , Castração , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Neoplasias de Próstata Resistentes à Castração/genética , Neoplasias de Próstata Resistentes à Castração/patologia
13.
J Biol Chem ; 286(10): 8117-8127, 2011 Mar 11.
Artigo em Inglês | MEDLINE | ID: mdl-21212260

RESUMO

D-type cyclins regulate cellular outcomes in part through cyclin-dependent, kinase-independent mechanisms that modify transcription factor action, and recent in vivo studies showed that cyclin D1 associates with a large number of transcriptional regulators in cells of the retina and breast. Given the frequency of cyclin D1 alterations in cancer, it is imperative to delineate the molecular mechanisms by which cyclin D1 controls key transcription factor networks in human disease. Prostate cancer was used as a paradigm because this tumor type is reliant at all stages of the disease on androgen receptor (AR) signaling, and cyclin D1 has been shown to negatively modulate AR-dependent expression of prostate-specific antigen (KLK3/PSA). Strategies were employed to control cyclin D1 expression under conditions of hormone depletion, and the effect of cyclin D1 on subsequent androgen-dependent gene expression was determined using unbiased gene expression profiling. Modulating cyclin D1 conferred widespread effects on androgen signaling and revealed cyclin D1 to be a selective effector of hormone action. A subset of androgen-induced target genes, known to be directly regulated by AR, was strongly suppressed by cyclin D1. Analyses of AR occupancy at target gene regulatory loci of clinical relevance demonstrated that cyclin D1 limits AR residence after hormone stimulation. Together, these findings reveal a new function for cyclin D1 in controlling hormone-dependent transcriptional outcomes and demonstrate a pervasive role for cyclin D1 in regulating transcription factor dynamics.


Assuntos
Androgênios/metabolismo , Ciclina D1/metabolismo , Regulação Neoplásica da Expressão Gênica , Neoplasias da Próstata/metabolismo , Receptores Androgênicos/metabolismo , Transdução de Sinais , Linhagem Celular Tumoral , Ciclina D1/genética , Loci Gênicos/genética , Humanos , Calicreínas/biossíntese , Calicreínas/genética , Masculino , Antígeno Prostático Específico/biossíntese , Antígeno Prostático Específico/genética , Neoplasias da Próstata/genética , Receptores Androgênicos/genética
14.
Breast Cancer Res ; 14(5): R130, 2012 Oct 04.
Artigo em Inglês | MEDLINE | ID: mdl-23036105

RESUMO

INTRODUCTION: Signal transducer and activator of transcripton-5a (Stat5a) and its close homologue, Stat5b, mediate key physiological effects of prolactin and growth hormone in mammary glands. In breast cancer, loss of nuclear localized and tyrosine phosphorylated Stat5a/b is associated with poor prognosis and increased risk of antiestrogen therapy failure. Here we quantify for the first time levels of Stat5a and Stat5b over breast cancer progression, and explore their potential association with clinical outcome. METHODS: Stat5a and Stat5b protein levels were quantified in situ in breast-cancer progression material. Stat5a and Stat5b transcript levels in breast cancer were correlated with clinical outcome in 936 patients. Stat5a protein was further quantified in four archival cohorts totaling 686 patients with clinical outcome data by using multivariate models. RESULTS: Protein levels of Stat5a but not Stat5b were reduced in primary breast cancer and lymph node metastases compared with normal epithelia. Low tumor levels of Stat5a but not Stat5b mRNA were associated with poor prognosis. Experimentally, only limited overlap between Stat5a- and Stat5b-modulated genes was found. In two cohorts of therapy-naïve, node-negative breast cancer patients, low nuclear Stat5a protein levels were an independent marker of poor prognosis. Multivariate analysis of two cohorts treated with antiestrogen monotherapy revealed that low nuclear Stat5a levels were associated with a more than fourfold risk of unfavorable outcome. CONCLUSIONS: Loss of Stat5a represents a new independent marker of poor prognosis in node-negative breast cancer and may be a predictor of response to antiestrogen therapy if validated in randomized clinical trials.


Assuntos
Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Fator de Transcrição STAT5/metabolismo , Adulto , Idoso , Neoplasias da Mama/mortalidade , Neoplasias da Mama/terapia , Núcleo Celular/metabolismo , Terapia Combinada , Progressão da Doença , Feminino , Humanos , Pessoa de Meia-Idade , Gradação de Tumores , Recidiva Local de Neoplasia , Estadiamento de Neoplasias , Avaliação de Resultados da Assistência ao Paciente , Fosforilação , Prognóstico , Transporte Proteico , Resultado do Tratamento , Carga Tumoral
15.
Gastroenterology ; 141(4): 1439-50, 2011 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-21704587

RESUMO

BACKGROUND & AIMS: The tumor suppressors retinoblastoma (RB) and p53 are important regulators of the cell cycle. Although human cancer cells inactivate RB and p53 by many mechanisms, the cooperative roles of these proteins in tumorigenesis are complex and tissue specific. We analyzed the cooperation of RB and p53 in liver development and pathogenesis of hepatocellular carcinoma. METHODS: Spontaneous and carcinogen-induced (diethylnitrosamine) tumorigenesis were studied in mice with liver-specific deletions of Rb and/or p53 (Rbf/f;albcre+, p53f/f;albcre+ and Rbf/f; p53f/f;albcre+ mice). Genotype, histologic, immunohistochemical, microarray, quantitative polymerase chain reaction, immunoblot, and comparative genomic hybridization analyses were performed using normal and tumor samples. Comparative microarray analyses were performed against publicly available human microarray data sets. RESULTS: Deletion of RB and p53 from livers of mice deregulated the transcriptional programs associated with human disease. These changes were not sufficient for spontaneous tumorigenesis; potent quiescence mechanisms compensated for loss of these tumor suppressors. In response to hepatocarcinogen-induced damage, distinct and cooperative roles of RB and p53 were revealed; their loss affected cell cycle control, checkpoint response, and genome stability. In damaged tissue, combined loss of RB and p53 resulted in early lesion formation, aggressive tumor progression, and gene expression signatures and histologic characteristics of advanced human hepatocellular carcinoma. CONCLUSIONS: The effects RB and p53 loss are determined by the tissue environment; cell stresses that promote aggressive disease reveal the functions of these tumor suppressors.


Assuntos
Carcinoma Hepatocelular/prevenção & controle , Neoplasias Hepáticas Experimentais/prevenção & controle , Fígado/metabolismo , Proteína do Retinoblastoma/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Animais , Carcinoma Hepatocelular/induzido quimicamente , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patologia , Ciclo Celular , Proliferação de Células , Aberrações Cromossômicas , Hibridização Genômica Comparativa , Dietilnitrosamina , Perfilação da Expressão Gênica/métodos , Regulação Neoplásica da Expressão Gênica , Instabilidade Genômica , Genótipo , Humanos , Immunoblotting , Imuno-Histoquímica , Fígado/patologia , Neoplasias Hepáticas Experimentais/induzido quimicamente , Neoplasias Hepáticas Experimentais/metabolismo , Neoplasias Hepáticas Experimentais/patologia , Camundongos , Camundongos da Linhagem 129 , Camundongos Knockout , Análise de Sequência com Séries de Oligonucleotídeos , Fenótipo , Proteína do Retinoblastoma/deficiência , Proteína do Retinoblastoma/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de Tempo , Transcrição Gênica , Proteína Supressora de Tumor p53/deficiência , Proteína Supressora de Tumor p53/genética
16.
Breast Cancer Res Treat ; 133(3): 1009-24, 2012 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-22134623

RESUMO

Ductal carcinoma in situ (DCIS) is a precursor lesion that can gives rise to invasive breast cancer (IBC). It has been proposed that both the nature of the lesion and the tumor microenvironment play key roles in progression to IBC. Here, laser capture microdissected tissue from pure DCIS and pure IBC were employed to define key gene expression profiles in either the epithelial or stromal compartment associated with disease progression. Each tissue had distinct gene expression profiles, and a DCIS/IBC classifier accurately distinguished DCIS versus IBC in multiple independent data sets. However, contrary to other studies that profiled DCIS associated with invasive disease, we found that the most significant alterations in gene expression were observed in the epithelial compartment rather than in the stroma. In particular, genes associated with epithelial-to-mesenchymal transition and myoepithelial cell-specific genes were enriched in invasive cancer relative to pure DCIS. Such alterations in transcript levels were associated with all subtypes of breast cancer, but were particularly indicative of poor outcome in ER-negative breast cancer. Together, these studies indicate that lesion-specific differences in gene expression associated with invasive phenotype are particularly relevant in the progression of DCIS to invasive breast cancer.


Assuntos
Neoplasias da Mama/genética , Carcinoma Ductal/genética , Carcinoma Intraductal não Infiltrante/genética , Transição Epitelial-Mesenquimal/genética , Regulação Neoplásica da Expressão Gênica , Neoplasias da Mama/mortalidade , Neoplasias da Mama/patologia , Carcinoma Ductal/mortalidade , Carcinoma Ductal/patologia , Carcinoma Intraductal não Infiltrante/mortalidade , Carcinoma Intraductal não Infiltrante/patologia , Análise por Conglomerados , Progressão da Doença , Feminino , Perfilação da Expressão Gênica , Humanos , Receptores de Estrogênio/deficiência , Reprodutibilidade dos Testes , Células Estromais/metabolismo , Células Estromais/patologia , Análise de Sobrevida
17.
Am J Pathol ; 179(3): 1171-8, 2011 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-21756866

RESUMO

The prevalence of ductal carcinoma in situ (DCIS) diagnoses has significantly increased as a result of active radiographic screening. Surgical resection and hormone and radiation therapies are effective treatments, but not all DCIS will progress to invasive breast cancer. Therefore, markers are needed to define tumors at low risk of recurrence and progression that can be treated by surgery alone rather than by adjuvant therapies. Initial analyses indicate that retinoblastoma (RB)-pathway perturbations occur at high frequency in DCIS and mirror the molecular alterations observed in invasive breast cancer. Particularly, the elevated expression of p16ink4a in DCIS was associated with loss of RB function and estrogen receptor-negative biology. Furthermore, high expression of p16ink4a in conjunction with Ki-67 was associated with increased risk of DCIS recurrence and progression to invasive disease in multivariate analyses. These data are consistent with a functional role for RB in modulating the invasive behavior of mammary epithelial cells. The tissue microenvironment is particularly relevant to the behavior of DCIS, and, surprisingly, elevated expression of p16ink4a in nonproliferative stroma was observed in a substantial fraction of cases. In this tissue compartment, p16ink4a expression was strongly associated with disease recurrence, independent of standard histopathologic features. Together, the data herein describe dual aspects of RB-pathway biology that are associated with disease recurrence through the epithelial or stromal compartment of DCIS.


Assuntos
Neoplasias da Mama/genética , Carcinoma Intraductal não Infiltrante/genética , Genes do Retinoblastoma/genética , Recidiva Local de Neoplasia/genética , Adulto , Idoso , Neoplasias da Mama/mortalidade , Neoplasias da Mama/cirurgia , Carcinoma Intraductal não Infiltrante/mortalidade , Carcinoma Intraductal não Infiltrante/cirurgia , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Feminino , Humanos , Estimativa de Kaplan-Meier , Antígeno Ki-67/metabolismo , Análise em Microsséries , Pessoa de Meia-Idade , Células Tumorais Cultivadas , Microambiente Tumoral/fisiologia
18.
Cell Mol Gastroenterol Hepatol ; 13(4): 1276-1296, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-34954189

RESUMO

BACKGROUND & AIMS: Sporadic colorectal cancers arise from initiating mutations in APC, producing oncogenic ß-catenin/TCF-dependent transcriptional reprogramming. Similarly, the tumor suppressor axis regulated by the intestinal epithelial receptor GUCY2C is among the earliest pathways silenced in tumorigenesis. Retention of the receptor, but loss of its paracrine ligands, guanylin and uroguanylin, is an evolutionarily conserved feature of colorectal tumors, arising in the earliest dysplastic lesions. Here, we examined a mechanism of GUCY2C ligand transcriptional silencing by ß-catenin/TCF signaling. METHODS: We performed RNA sequencing analysis of 4 unique conditional human colon cancer cell models of ß-catenin/TCF signaling to map the core Wnt-transcriptional program. We then performed a comparative analysis of orthogonal approaches, including luciferase reporters, chromatin immunoprecipitation sequencing, CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats) knockout, and CRISPR epigenome editing, which were cross-validated with human tissue chromatin immunoprecipitation sequencing datasets, to identify functional gene enhancers mediating GUCY2C ligand loss. RESULTS: RNA sequencing analyses reveal the GUCY2C hormones as 2 of the most sensitive targets of ß-catenin/TCF signaling, reflecting transcriptional repression. The GUCY2C hormones share an insulated genomic locus containing a novel locus control region upstream of the guanylin promoter that mediates the coordinated silencing of both genes. Targeting this region with CRISPR epigenome editing reconstituted GUCY2C ligand expression, overcoming gene inactivation by mutant ß-catenin/TCF signaling. CONCLUSIONS: These studies reveal DNA elements regulating corepression of GUCY2C ligand transcription by ß-catenin/TCF signaling, reflecting a novel pathophysiological step in tumorigenesis. They offer unique genomic strategies that could reestablish hormone expression in the context of canonical oncogenic mutations to reconstitute the GUCY2C axis and oppose transformation.


Assuntos
Neoplasias Colorretais , beta Catenina , Carcinogênese/genética , Cateninas/genética , Cateninas/metabolismo , Neoplasias Colorretais/patologia , Humanos , Ligantes , Região de Controle de Locus Gênico , Receptores de Enterotoxina/genética , Receptores de Enterotoxina/metabolismo , Fatores de Transcrição TCF/metabolismo , beta Catenina/genética , beta Catenina/metabolismo
19.
Cancer Res ; 82(3): 458-471, 2022 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-34903608

RESUMO

Despite treatment with intensive chemotherapy, acute myelogenous leukemia (AML) remains an aggressive malignancy with a dismal outcome in most patients. We found that AML cells exhibit an unusually rapid accumulation of the repressive histone mark H3K27me3 on nascent DNA. In cell lines, primary cells and xenograft mouse models, inhibition of the H3K27 histone methyltransferase EZH2 to decondense the H3K27me3-marked chromatin of AML cells enhanced chromatin accessibility and chemotherapy-induced DNA damage, apoptosis, and leukemia suppression. These effects were further promoted when chromatin decondensation of AML cells was induced upon S-phase entry after release from a transient G1 arrest mediated by CDK4/6 inhibition. In the p53-null KG-1 and THP-1 AML cell lines, EZH2 inhibitor and doxorubicin cotreatment induced transcriptional reprogramming that was, in part, dependent on derepression of H3K27me3-marked gene promoters and led to increased expression of cell death-promoting and growth-inhibitory genes.In conclusion, decondensing H3K27me3-marked chromatin by EZH2 inhibition represents a promising approach to improve the efficacy of DNA-damaging cytotoxic agents in patients with AML. This strategy might allow for a lowering of chemotherapy doses, with a consequent reduction of treatment-related side effects in elderly patients with AML or those with significant comorbidities. SIGNIFICANCE: Pharmacological inhibition of EZH2 renders DNA of AML cells more accessible to cytotoxic agents, facilitating leukemia suppression with reduced doses of chemotherapy.See related commentary by Adema and Colla, p. 359.


Assuntos
Cromatina/metabolismo , Histonas/metabolismo , Leucemia Mieloide Aguda/genética , Animais , Humanos , Camundongos
20.
BMC Genomics ; 12: 464, 2011 Sep 26.
Artigo em Inglês | MEDLINE | ID: mdl-21943378

RESUMO

BACKGROUND: The creation of lymphoblastoid cell lines (LCLs) through Epstein-Barr virus (EBV) transformation of B-lymphocytes can result in a valuable biomaterial for cell biology research and a renewable source of DNA. While LCLs have been used extensively in cellular and genetic studies, the process of cell transformation and expansion during culturing may introduce genomic changes that may impact their use and the interpretation of subsequent genetic findings. RESULTS: We performed whole exome sequencing on a tetrad family using DNA derived from peripheral blood mononuclear cells (PBMCs) and LCLs from each individual. We generated over 4.7 GB of mappable sequence to a 125X read coverage per sample. An average of 19,354 genetic variants were identified. Comparison of the two DNA sources from each individual showed an average concordance rate of 95.69%. By lowering the variant calling parameters, the concordance rate between the paired samples increased to 99.82%. Sanger sequencing of a subset of the remaining discordant variants did confirm the presence of de novo mutations arising in LCLs. CONCLUSIONS: By varying software stringency parameters, we identified 99% concordance between DNA sequences derived from the two different sources from the same donors. These results suggest that LCLs are an appropriate representation of the genetic material of the donor and suggest that EBV transformation can result in low-level generation of de novo mutations. Therefore, use of PBMC or early passage EBV-transformed cells is recommended. These findings have broad-reaching implications, as there are thousands of LCLs in public biorepositories and individual laboratories.


Assuntos
Linfócitos B/virologia , Transformação Celular Viral , DNA/química , Herpesvirus Humano 4/fisiologia , Leucócitos Mononucleares/metabolismo , Linfócitos B/fisiologia , Doadores de Sangue , Linhagem Celular Transformada , Transformação Celular Viral/genética , DNA/metabolismo , Exoma/genética , Humanos , Análise de Sequência de DNA
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA